Tumor growth and migration are dependent on angiogenesis and lymphangiogenesis. Therefore, tumor angiogenic and lymphangiogenic endothelial cells have emerged as new targets for cancer therapeutics (Folkman, J. N Engl J Med 1971; 285:1182-1186, Witte M H et al. Lymphology. 1987; 20(4):257-66). Meanwhile, it has been shown that abnormal angiogenesis is also involved in a variety of diseases including obesity, diabetic retinopathy, persistent hyperplastic vitreous, psoriasis, allergic dermatitis, arthritis, arteriosclerosis, endometriosis, asthma, ascites, and peritoneal adhesion (Carmeliet P., Nature Medicine 2003; 9(6): 653-660).
Endostatin (ES) is a 20 kDa C-terminal fragment of collagen XVIII and is an endogenous inhibitor of angiogenesis and lymphangiogenesis. ES can inhibit the proliferation and migration of angiogenic and lymphangiogenic endothelial cells, and thus can impede the formation of new blood vessels and lymphatic vessels. It has been shown that recombinant endostatin can inhibit the growth and metastasis of various types of tumors without eliciting drug resistance in animal models (Folkman J. et al. Cell 1997; 88:277-285, Folkman J. et al. Nature 1997; 390:404-407, Zhuo W. et al, Journal of Pathology; 222:249-260). Recently, a recombinant human endostatin, Endu, containing an extra N-terminal (M)GGSHHHHH amino acid residues compared to the natural endostatin, has been widely used and tested in clinic for the treatment of multiple types of cancers, especially non-small cell lung cancer. It has been shown that ES can be internalized by activated angiogenic and lymphangiogenic endothelial cells. However, the specific mechanism remains unclear.
Epithelial Growth Factor Receptor (EGFR) is overexpressed on the surface of various types of tumor cells and is closely related to the proliferation and metastasis of tumor cells. Several monoclonal antibodies against EGFR have been developed as antitumor therapeutics. For example, Cetuximab is a monoclonal antibody of EGFR that specifically binds to cell surface EGFR of tumor cells. Cetuximab has been used for the treatment of certain cancers.
Nystatin (NT) is a polyene antibiotic that is used as a broad spectrum antifungal drug in human. It can be orally or topically applied, without any interaction with other drugs. NT has high efficacy against Candida, while other fungi including cryptococcus neoformans, aspergillus, mucor, microsporum, histoplasma capsulatum, blastomyces dermatitidis, dermatophytes are usually sensitive to NT as well. NT is also commonly used as a prophylactic treatment in patients prone to fungal infection, such as AIDS patients and patients undergoing chemotherapy. The mechanism of nystatin and its analogs (e.g., amphotericin B) is that they can bind with ergosterol on the fungus cell membrane, resulting in membrane perforation, potassium leakage, and subsequently the death of fungus. Since ergosterol is a unique lipid composition of fungus cell membrane, NT and amphotericin B will not affect eukaryotic cell membrane when applied to human or animals.
NT can influence the cholesterol-dependent endocytosis of cells and has been used as a specific inhibitor of lipid raft/caveolae-dependent endocytic pathway. Since cholesterol is required for lipid raft/caveolae-dependent endocytic pathway, NT can bind with cell surface membrane cholesterol, thus inhibiting this pathway. Amphotericin B, methyl-β-cyclodextrin, and filipin have a similar function to that of NT and can also inhibit lipid raft/caveolae-dependent endocytic pathway by affecting cholesterol. Amphotericin B (AMB) has the same function as NT and is used for the treatment of cryptococcus and aspergillus infection.
Wickstrom et al (2002, 2003) reported that in endothelial cells, ES can bind to integrin alpha5beta1 and caveolin (the structural protein of caveolae) in lipid rafts, indicating the correlation between ES and lipid raft (Wickstrom et al, Cancer Res. 2002; 62: 5580-9, Wickstrom et al, J. Biol. Chem. 2003; 278(39): 37895-37901). In addition, Dixelius et al (2000), Shi et al (2007), and Zhuo et al (2010) respectively reported that ES can be specifically internalized in angiogenic and lymphangiogenic endothelial cells (Dixelius et al, Blood 2000, 95, 3403-3411; Shi et al, Blood 2007, 110, 2899-2906, Zhuo W. et al, Journal of Pathology; 222:249-260).
In 2002, Pike et al and Roepstorff et al found that the cell surface binding of epithelial growth factor (EGF), the ligand of EGFR, can be increased by inhibition of cholesterol and lipid rafts (Pike et al, Biochemistry 2002; 41: 10315-22, Roepstorff et al, J Biol Chem 2002; 277: 18954-60).
Kojic et al (2008) reported that tumor cells can internalize AMF/PGI through the lipid raft/caveolae-dependent endocytic pathway. In addition, this endocytic process is cell-type specific, indicating that lipid raft/caveolae-dependent endocytosis might provide a drug delivery route specifically targeting tumor cells (Kojic et al, PLoS ONE 2008; 3: e3597). Migalovich et al (2009) reported that NT increases the uptake of daidzein-BSA by ovarian cancer cells through regulating lipid raft/caveolae-dependent endocytosis. Daidzein-BSA is a potential tumor imaging agent. By increasing the uptake of daidzein-BSA by tumor cells, NT may improve the effects of this tumor imaging agent in tumor imaging (Migalovich et al, Cancer Res. 2009; 69: 5610-5617). In conclusion, lipid raft/caveolae-dependent endocytosis may be involved in the uptake of many important molecules including cytokines, exogenous proteins, and even therapeutic compounds. Regulating and manipulating such endocytic procedures at a cellular level may influence the uptake of these substances in a subject, which may find a potential application for drug delivery.